The invention relates to apparatus and a method for mixing two liquids within a tip on an aspirating probe, to ensure a reaction between the liquids.
It is known from U.S. Pat. Nos. 5,773,305 and 5,174,162 to mix a fluid sample such as blood and a diluent, inside a probe tip by first aspirating both liquids into the tip, and then drawing said liquids further up into the tip into a mixing chamber having an enlarged inside diameter compared to the rest of the tip. The mixing can be achieved, for example, by reciprocating the mass of liquids up and down numerous times.
In the examples shown in U.S. Pat. No. 5,773,305, the liquids are retained in the enlarged chamber and simply sloshed back and forth in that chamber to achieve mixing. FIG. 3 thereof makes it clear that simply aspirating the liquids into the enlarged chamber past a step discontinuity created by the enlarged inside diameter, is ineffective in creating a mixture. That is, a single movement past the step discontinuity is shown as not mixing the fluids homogeneously. An air bubble can also be included between the liquids when first aspirated. Cross-over contamination between bodies of liquid being aspirated is preferably prevented by ejecting an inert oil shield around the outside of the tip, FIGS. 7 through 11 thereof.
Such a construction is generally equivalent to transferring two liquids from a pipette into a larger diameter container (the mixing chamber) and attempting mixing by sloshing the liquids vertically within the container. Although mixing can occur in such a fashion for relatively large volumes, it is not as effective for small volumes, e.g., volumes that total 100 to 600 microliters. That is, in a constant diameter channel, inertial mixing is reduced if the volumes are small, as here. It is this phenomenon that requires the movement of the liquids back and forth in the mixing chamber, as much as 20 times, to achieve homogeneous mixing. Such reiterations of the mix step are time-consuming, and beg for an improvement.
Furthermore, it is not the case that cross-contamination is preventable only by using such an oil shield. That is, in some cases, the first-aspirated liquid can be removed from the tip simply be repeated washing with a diluent, or by wiping. In any event, should washing prove to be unsatisfactory, there has been a need for a more reliable method of preventing contamination than by using the oil shield. (The oil shield is not guaranteed to form completely around the tip just because a plurality of dispensing nozzles are disposed about the circumference of the exterior of the tip.) Furthermore, some proteins can destroy the shield effect of the oil.
In the examples of U.S. Pat. No. 5,174,162, all the liquids to be mixed are moved completely into the enlarged mixing chamber, completely out of the chamber, then back into it, and so forth. The sharp transition at surface 15 causes turbulent mixing, 16, FIG. 2 thereof. This is a more efficient mixing method than that of the ""305 patent. Nevertheless, there are improvements that are needed in such a mixing system as described in the ""162 patent. For example, no optimization is described for the geometry of FIG. 2. Nothing is described regarding any use of air bubbles to separate the liquids as they are aspirated. As noted however in the ""305 patent, such an air bubble provides an effective prevention against cross-contamination. Yet, any air bubble must be rapidly eliminated during mixing.
Furthermore, the ""162 patent is notably deficient in any teaching to prevent cross-contamination when aspirating liquid 6 immediately after liquid 4, between the two liquids within the bulk container of liquid 6. Although the oil shield of the ""305 patent might seem to be applicable to the probe of the ""162 patent as well, such a shield has disadvantages as noted above. Alternative protection methods against cross-contamination, besides the oil-shield method, are thus desirable.
Yet another disadvantage of the teachings of the ""162 patent is that when the two disparate liquids are moved back and forth across the boundary 15, unmixed xe2x80x9ctailsxe2x80x9d of one or both liquids can be left behind as coatings on either the enlarged chamber or the narrower intake portion. Such residual tails do not get mixed when the main body of liquids is moved across boundary 15, so that the tails are undesirable.
Thus, although substantial development has already occurred in probes designed to mix two liquids entirely with the probe, there remains the need for improvements.
We have devised a mixing method and a probe tip for doing the mixing therein, that provide the above-noted needed improvements.
More specifically, in accord with one aspect of the invention, there is provided a method of mixing a plurality of liquids, comprising the steps of:
a) providing a probe tip with an internal cavity having a plurality of different inside diameters;
b) providing by aspiration a plurality of liquids inside a portion of the probe tip;
c) moving at least most of said liquids back and forth at least several times between a part of said cavity with a smaller inside diameter and a part with a larger inside diameter, said larger and smaller diameters being sufficient to provide a sufficient rotation of liquid as it moves between diameters to cause mixing of said liquids;
the improvement wherein the capillary number resulting from the mixing in step c) does not exceed about 0.01, the capillary number being defined as the ratio of liquid velocity times viscosity and surface tension, so that any tails formed during the mixing step c) are minimized.
In accord with another aspect of the invention, there is provided a method of mixing a plurality of liquids comprising the steps of a) through c) listed above, wherein the improvement comprises that the cavity parts comprise two separate but matable tip portions, and the method further includes the step of mounting a tip portion of one of the inside diameters onto the tip portion of the other inside diameter in-between aspiration of liquids, such that carry-over contamination between liquids is prevented.
In accord with still another aspect of the invention, there is provided a method of mixing a plurality of liquids comprising the steps of a) through c) listed above, wherein the improvement comprises the inside diameters each provide a cross-sectional flow-through area of the cavity part, and the cross-sectional flow-through area of the larger inside diameter is at least three times the cross-sectional flow through area of the smaller inside diameter, for maximum mixing efficiency.
In accord with yet another aspect of the invention, there is provided a method of mixing a plurality of liquids comprising the steps of a) through c) listed above, wherein the improvement comprises the larger inside diameter being obtained by i) selecting as a first tip portion a tapered tip at least a portion of which has an inside diameter that is much larger than the smaller inside diameter of the probe tip, and ii) joining the tapered tip to the probe tip having the smaller inside diameter.
In accord with yet another aspect of the invention, there is provided a method of mixing a plurality of liquids comprising the steps of a) through c) listed above, wherein the improvement comprises providing a total amount of liquid in step b) such that if all liquid is moved into the part with the larger inside diameter, the larger inside diameter is greater than the height of the total liquid, but less than twice the height of the total liquid, so that mixing as per step c) is maximized.
In accord with yet another aspect of the invention, there is provided a method of mixing a plurality of liquids comprising the steps of a) through c) listed above, wherein the improvement comprises moving in the step c) at least most of the liquids back and forth at least between the cavity part with the smaller inside diameter and a part of the cavity of a larger inside diameter located at opposite ends of the cavity part of the smaller inside diameter, so that mixing efficiency is enhanced by rotation of the liquid as it moves past the opposite ends, rather than a single end of the smaller inside diameter cavity part.
In accord with yet another aspect of the invention, there is provided a method of mixing a plurality of liquids comprising the steps of a) through c) listed above, wherein the improvement comprises moving in the step c) at least most of the liquids back and forth at least between the cavity part with the smaller inside diameter and a part of the cavity of a larger inside diameter located at opposite ends of the cavity part of the smaller inside diameter, so that mixing efficiency is enhanced by rotation of the liquid as it moves past the opposite ends, rather than a single end of the smaller inside diameter cavity part.
In accord with yet another aspect of the invention, there is provided a probe tip for mixing liquids within the tip after aspiration of the liquids therein to, the tip comprising
a wall defining 3 connected cavities of unequal inside diameters one of the compartments being sandwiched as a middle compartment between the other two which form end compartments, each two adjacent cavities being connected by a transition zone wall and the inside diameters being sufficiently unequal in the adjacent 2 cavities as to cause rotational mixing of liquids as they move past the transition zone wall,
wherein the transition zone of the one cavity is formed by a variance of the inside diameter that increases in value as the middlemost cavity is transited outward into either of the other two end cavities.
In accordance with yet another aspect of the invention, there is provided a method of determining the strength of an agglutination reaction within a hollow container comprising walls capable of transmitting light at certain predetermined wavelengths, comprising the steps of:
a) providing a mixture of a sample and an agglutinating reagent within a first cavity of the container, the cavity having a first inside diameter,
b) transferring the mixture to a second cavity having a second inside diameter substantially smaller than the first inside diameter,
c) scanning the liquid within the second cavity during the step b) with a beam of light at the predetermined wavelengths, the 10% portion being that portion closest to the first cavity;
d) after the scanning step c), detecting the amount of light absorbed within or scattered by the 10% portion by the beam,
e) transferring the mixture back into the first cavity,
f) repeating steps b)-d) at least once until some agglutinated material has separated from non-agglutinated material, and
g) calculating the amount of agglutination from the absorbance or scattering detected in step d).
In accordance with yet another aspect of the invention, there is provided a method of agglutinating blood cells in whole blood, comprising the steps of
a) aspirating whole blood into a disposable tip mounted on a probe, said tip having at least two portions with significantly different inside diameters, connected to each other by a transition zone,
b) aspirating into the same tip thereafter, an agglutinating reagent, and
c) moving said blood and reagent back and forth as a total liquid, first entirely into one of said portions and then entirely into the other of said portions, a sufficient number of times so as to cause coagulation of the cells of the whole blood, and then subsequent separation of plasma from the coagulated cells.
As used herein, xe2x80x9cprobe tipxe2x80x9d or xe2x80x9cprobe tip portionxe2x80x9d means any vessel, disposable or not, into which liquid can be aspirated, mountable on an aspirating probe, that comprises the features noted, namely an orifice, an interior chamber spaced from the orifice, and a passageway connecting the orifice and the chamber. Thus, the tip or tip portion can be a conventional disposable tip such as is shown in U.S. Pat. No. 4,347,875 by Columbus, or even a cup or well with an orifice in the bottom, such as the cup shown in U.S. Pat. No. 5,441,895 but with an orifice in the bottom. The tip can be one integral piece or provided in several portions.
Accordingly, it is an advantageous feature of the invention that more rapid mixing of two liquids aspirated into the tip, takes place within the tip than occurs with conventional devices.
It is a related advantageous feature of the invention that no additional device is needed beyond the tip that is used anyway for aspiration, to provide mixing.
It is another advantageous feature of the invention that, in some embodiments, carry-over contamination between liquids aspirated is preventable by an inexpensive mechanical device that is less time consuming than repeated washing.
A related advantage of the aforesaid mechanical device for preventing carry-over contamination, is that it renders the tip of the invention more manufacturable.
Other advantageous features will become apparent upon reference to the Detailed Description of the Embodiments, when read in light of the attached drawings.